The use of microfluidic technology has been proposed for a number of analytical chemical and biochemical operations. This technology allows one to perform chemical and biochemical reactions, macromolecular separations, and the like, that range from the simple to the relatively complex, in easily automated, high-throughput, low-volume systems. Further information about microfluidic devices and systems is presented in U.S. Pat. No. 6,534,013 to Kennedy, issued Mar. 18, 2003, and incorporated in its entirety herein by reference.
As used herein, the term “microfluidic,” or the term “microscale” when used to describe a fluidic element, such as a passage, chamber or conduit, generally refers to one or more fluid passages, chambers or conduits which have at least one internal cross-sectional dimension, e.g., depth or width, of between about 0.1 μm and 500 μm. In the devices of the present invention, the microscale channels preferably have at least one cross-sectional dimension between about 0.1 μm and 200 μm, more preferably between about 0.1 μm and 100 μm, and often between about 0.1 μm and 20 μm.
In general, microfluidic systems include a microfluidic device, or chip, that has networks of integrated submicron channels in which materials are transported, mixed, separated, and detected. Microfluidic systems typically also contain components that provide fluid driving forces to the chip and that detect signals emanating from the chip.
Microfluidic chips may be fabricated from a number of different materials, including glass or polymeric materials. An example of a commercially available microfluidic chip is the DNA LabChip® manufactured by Caliper Life Sciences, Inc. of Hopkinton, Mass., and used with the Agilent 2100 Bioanalyzer system manufactured by Agilent Technologies, Inc. of Palo Alto, Calif. The chip has two major components: a working part made of glass, and a plastic caddy or mount bonded to the working part. The working part contains microfluidic channels in its interior, and wells on its exterior that provide access to the microfluidic channels. The working part is typically fabricated by bonding together two or more planar substrate layers. The microfluidic channels in the working part are formed when one planar substrate encloses grooves formed on another planar substrate. The mount protects the working part of the chip, and provides for easier handling of the chip by a user. The increased ease of handling partially results from the fact that the mount is larger than the working part of the device, which in many cases is too small and thin to be easily handled. The mount may be fabricated from any suitable polymeric material, such as an acrylic or thermoplastic. The glass working part is typically bonded to the polymeric mount using a UV-cured adhesive. Reservoirs in the mount provide access to the wells on the working part of the chip. The reservoirs hold much greater volumes of material than the wells in the working part, thus providing an interface between the macro-environment of the user and the microenvironment of the wells and channels of the microfluidic device.
This type of microfluidic chip is a “planar” chip. In a planar chip, the only access to the microchannels in the chip is through the reservoirs in the caddy and in-turn through the wells in the working part. Another type of microfluidic chip is a “sipper” chip, which has a small tube or capillary (the “sipper”) extending from the chip through which fluids stored outside the chip can be directed into the microfluidic channels in the chip. Typical sipper chips have between one and twelve sippers. In use, the sipper is placed in a receptacle having sample material and minute quantities of the sample material are introduced, or “sipped” through the capillary tube to the microfluidic channels of the chip. This sipping process can be repeated to introduce any number of different sample materials into the chip. Sippers make it easier to carry out high-throughput analysis of numerous samples on a single microfluidic chip.
Western blot electrophoresis assays have been developed to detect specific proteins in a sample. The process can be divided into three parts: protein separation, sample transfer, and immunoassay. In protein separation, mechanical and/or chemical techniques are applied to a sample, such as a tissue sample, to expose proteins. The proteins are then separated with gel electrophoresis in which the speed of movement of the different proteins through the gel under a differential voltage is governed by the molecular weight of the individual proteins. In sample transfer, the separated proteins are moved from within the gel onto a membrane in a process called electroblotting, which uses electric current to move the proteins. In immunoassay, a primary antibody is attached to target proteins on the membrane, a secondary antibody is attached to the primary antibody, and a light emitter reacts with the secondary antibody to produce light at each of the target proteins. Detection of the light provides identification and quantification of the target proteins.
Although the current method of Western blot electrophoresis assay provides valuable results, the current method has a number of problems. The current method is a labor-intensive process, performed manually and requiring gel plates and special membrane paper to transfer the separated proteins. The manual nature of the process increases the cost and limits the number of samples which can be tested. A typical Western analysis requires between 8 and 24 hours of monitored operation, with almost half requiring hands-on, manual operation.
It would be desirable to have methods and systems of microfluidic immunoassay using magnetic beads that would overcome the above disadvantages.